Proteins and other large biological molecules, including DNA, may be separated for analysis using electrophoresis techniques. One particularly important application of these techniques is the sequencing of the DNA and RNA molecules. In performing an electrophoresis separation of such molecules, a gel is formed between two non conducting plates, such as glass, to form a thin sheet of gel between the glass surfaces. The gel should be as uniform in thickness and other properties as possible. The surfaces at either end of the gel are each connected to a buffer reservoir which serve as electrodes. A potential is applied across the gel by connecting each reservoir to opposite polarities of a voltage source.
The collection of molecules to be sorted is placed at the negative electrode end of the gel, usually in pre-formed wells. The molecules are negatively charged, and the electric field in the gel reacts with the charge on the molecules to provide a force propelling the molecules through the gel towards the positive electrode. Smaller molecules have less resistance to traveling through the gel than larger molecules, resulting in a separation and sorting of the molecules by size as they migrate through the gel.
Synthesis of fractional lengths of DNA (or RNA) molecules by means of enzymes allows the molecule to be analyzed according to the sequence of bases making up such a molecule. Techniques for doing this type of analysis are well known. See for example Sanger, F., S. Nicklen & A.R. Coulson 1977 Proc. Natl. Acad. Sci. USA 74: 5463-5467; Prescott L. Deininger Analytical Biochemistry 135, 247 263 (1983); Garoff, H., & Ansorge, W. (1981) Anal. Biochem. 115, 450 457; and Biggin, M.D., T.J. Gibson & A.F. Hong 1983 Proc. Natl. Acad. Sci. USA 80: 3963-3965.
One method employed to view the electrophoretic separation of macromolecules in a gel involves radio-labelling the molecules. Typically, the gel used for electrophoresis is removed from its cassette and placed alongside a photographic medium which is exposed by the radioactive emissions of the radio-labelled molecules. Developing the emulsion produces a series of stripes representative of the position of each set of macromolecules in the case of sequencing their relative locations can be used by the investigator to derive the base relative locations sequence directly. This type of analysis requires a high degree of training, is time consuming, tedious and very prone to transcription errors. Automatic detection methods have been proposed in which detectors responsive to the radioactive emission are used to automatically perform DNA sequencing. However, due to variations from the ideal operation of the electrophoresis gel cassette, automatic systems have not performed well.
A system which is capable of performing an electrophoresis separation of macromolecules in such a way that relatively simple detection apparatus can reliably and accurately determine their position would have great advantages. Additionally, current methods for assembling gel cassettes for performing electrophoresis require great skill and considerable time. A gel cassette which could be assembled and loaded with gel by a relatively unskilled technician while retaining the above mentioned features would greatly aid in performing electrophoresis analysis of biological macromolecules.